Fig. 1

a Differential expression from RNA sequencing of embryos subjected to Keap1-hydroxynonenylation by Z-REX. Statistically-significant differentially-expressed (SDE) Nrf2-driven AR genes marked with green dots; SDE immunity-related genes with red dots; all other SDE genes with blue dots; and non-SDE genes with gray dots. b qRT-PCR analysis validated suppression of immunity-related genes in zebrafish embryos following Z-REX-assisted Keap1-specific hydroxynonenylation. P values were calculated with ANOVA and Tukey’s multiple comparisons test. c Same as in (a) except Z-REX-mediated modification of Keap1 was executed with a different electrophile, HDE, of similar Keap1-modification efficiency⁵³ and ability to upregulate AR genes [green dots]. Gray dashed lines mark genes not SDE in this comparison. See also Supplementary Table 1 and Supplementary Data 1. d Z-REX-mediated Keap1-hydroxynonenylation, but not Z-REX-technical controls (Supplementary Fig. 1a), caused depletion of neutrophil count in Tg(lyz:TagRFP), in which neutrophils are labeled with TagRFP. Scale bars, 500 μm. e Same as in (d) in fish expressing Halo-P2A-Keap1, which cannot undergo Keap1-hydroxynonenylation by Z-REX (Supplementary Fig. 1a). Scale bars, 500 μm. f Quantitation of neutrophil levels in Tg(lyz:TagRFP) following Z-REX (against all technical controls) using photocaged Z-REX probes, Ht-PreH(D)NE, delivering HN(D)E. Note: signal-to-noise in these experiments (d–f) is 6:1, so gene expression changes rendering signal below detection levels cannot explain this loss of neutrophils. Tukey’s multiple comparisons test was used to calculate corrected p values. g Similar experiment as in (d) in fish expressing either Halo-TEV-Keap1 or Halo-TEV-Keap1^{C151S&C273W&C288E} (‘Halo-TEV-Keap1 mutant’) which cannot undergo Keap1-hydroxynonenylation. Scale bars, 500 μm. h Quantitation of neutrophil levels in Tg(lyz:TagRFP) following Z-REX using photocaged Z-REX probes, Ht-PreHNE, delivering HNE (d and g). Inset: analysis of fold change (Z-REX/non-treated; see Fig. 2d) in neutrophil count. Note: for Halo-TEV-Keap1 set, the same data are presented in panels (f) and (h) for clarity. P values were calculated with two-tailed Student’s t-test. All data present mean ± SEM. All p values for differential expression in RNA-seq were calculated with CuffDiff. All sample sizes are listed in Supplementary Methods. Source data are provided as a Source data file.